Identification of a novel splice site mutation in the DNAAF4 gene of a Chinese patient with primary ciliary dyskinesia.

Primary ciliary dyskinesia (PCD) is a rare hereditary orphan condition that results in variable phenotypes, including infertility. About 50 gene variants are reported in the scientific literature to cause PCD, and among them, dynein axonemal assembly factor 4 ( DNAAF4 ) has been recently reported. DNAAF4 has been implicated in the preassembly of a multiunit dynein protein essential for the normal function of locomotory cilia as well as flagella. In the current study, a single patient belonging to a Chinese family was recruited, having been diagnosed with PCD and asthenoteratozoospermia. The affected individual was a 32-year-old male from a nonconsanguineous family. He also had abnormal spine structure and spinal cord bends at angles diagnosed with scoliosis. Medical reports, laboratory results, and imaging data were investigated. Whole-exome sequencing, Sanger sequencing, immunofluorescence analysis, hematoxylin-eosin staining, and in silico functional analysis, including protein modeling and docking studies, were used. The results identified DNAAF4 disease-related variants and confirmed their pathogenicity. Genetic analysis through whole-exome sequencing identified two pathogenic biallelic variants in the affected individual. The identified variants were a hemizygous splice site c.784-1G>A and heterozygous 20.1 Kb deletion at the DNAAF4 locus, resulting in a truncated and functionless DNAAF4 protein. Immunofluorescence analysis indicated that the inner dynein arm was not present in the sperm flagellum, and sperm morphological analysis revealed small sperm with twisted and curved flagella or lacking flagella. The current study found novel biallelic variants causing PCD and asthenoteratozoospermia, extending the range of DNAAF4 pathogenic variants in PCD and associated with the etiology of asthenoteratozoospermia. These findings will improve our understanding of the etiology of PCD.

Modulation of SIRT1 expression improves erectile function in aged rats.

Silent information regulator 2-related enzyme 1 (SIRT1) is an aging-related protein activated with aging. Herein, we evaluated the role of SIRT1 in aging-related erectile dysfunction. The expression of SIRT1 was modulated in aged Sprague-Dawley rats following intragastric administration of resveratrol (Res; 5 mg kg-1), niacinamide (NAM; 500 mg kg-1) or Res (5 mg kg-1) + tadalafil (Tad; phosphodiesterase-5 [PDE5] inhibitor; 5 mg kg-1) for 8 weeks. Then, we determined erectile function by the ratio of intracavernosal pressure (ICP)/mean systemic arterial pressure (MAP). Cavernosal tissues were extracted to evaluate histological changes, cell apoptosis, nitric oxide (NO)/cyclic guanosine monophosphate (cGMP), the superoxide dismutase (SOD)/3,4-methylenedioxyamphetamine (MDA) level, and the expression of SIRT1, p53, and forkhead box O3 (FOXO3a) using immunohistochemistry, terminal deoxynucleotidyl transferase (TdT)-mediated 2'-deoxyuridine 5'-triphosphate (dUTP) nick-end labeling (TUNEL), enzyme-linked immunosorbent assays, and western blot analysis. Compared with the control, Res treatment significantly improved erectile function, reflected by an increased content of smooth muscle and endothelium, NO/cGMP and SOD activity, and reduced cell apoptosis and MDA levels. The effect of Res was improved by adding Tad. In addition, the protein expression of SIRT1 was increased in the Res group, accompanied by decreased p53 and FOXO3a levels. In addition, inhibition of SIRT1 by NAM treatment resulted in adverse results compared with Res treatment. SIRT1 activation ameliorated aging-related erectile dysfunction, supporting the potential of SIRT1 as a target for erectile dysfunction treatment.

Bilateral is superior to unilateral varicocelectomy in infertile males with left clinical and right subclinical varicocele: a prospective randomized controlled study.

PURPOSE: The purpose of this study is to compare the effect of bilateral versus unilateral varicocelectomy on seminal response and spontaneous pregnancy rates in infertile male patients with left clinical and right subclinical varicocele. METHODS: A total of 358 infertile men with left clinical and right subclinical varicocele were randomized to group that underwent bilateral (n = 179) and group that underwent unilateral microsurgical subinguinal varicocelectomy (n = 179). Baseline data regarding male age, female partner age, grade of varicocele body mass index, bilateral testicular volume and serum follicle-stimulating hormone, luteinizing hormone, total testosterone levels and infertility duration and semen parameters were gathered. One year after the surgery, semen parameters including sperm volume, sperm concentration, normal sperm morphology, progressive motility and sperm DNA fragmentation index were recorded and any pregnancy was also documented via telephone calls and hospital visits. RESULTS: We found the baseline characteristics were comparable between the two groups. The seminal parameters had significant improvements 1 year postoperatively in both groups. However, the bilateral group showed significantly greater improvements than the unilateral group in sperm concentration, normal sperm morphology and progressive motility. Besides, the pregnancy rate was statistically higher in the bilateral group after the surgery (42.5 versus 26.0%, bilateral versus unilateral group). CONCLUSION: In conclusion, our study indicated that bilateral is superior to unilateral varicocelectomy in infertile males with left clinical and right subclinical varicocele, which is associated with greater improvements in sperm concentration, normal sperm morphology and progressive motility and spontaneous pregnancy rate after the surgery.

[Laparoscopic extraperitoneal high ligation of the spermatic vein for the treatment of varicocele].

OBJECTIVE: To investigate the effect of laparoscopic extraperitoneal (LSEP) high ligation of the spermatic vein in the treatment of varicocele. METHODS: We retrospectively analyzed the clinical data about 80 cases of varieocele, 48 treated by LSEP and the other 32 by laparoscopic transabdominal retroperitoneal (LSTR) high ligation of the spermatic vein. We recorded the semen parameters before and at 1, 3 and 6 months after surgery, intraoperative blood loss, operation time, postoperative complications, time of gastrointestinal function recovery and rate of pregnancy, followed by comparison of the data obtained between the two groups of patients. RESULTS: Semen parameters were remarkably improved in both the LSEP and LSTR groups of patients postoperatively as compared with the baseline (P <0.05), but with no statistically significant difference at different postoperative months (P >0.05). The intraoperative blood loss was less in the LSEP than in the LSTR group (ï¼»8.3 ± 5.2ï¼½ vs ï¼»9.1 ± 6.1ï¼½ ml, P >0.05), the operation time was shorter in the former than in the latter (ï¼»38.27 ± 9.23ï¼½ vs ï¼»43.46 ± 11.72ï¼½ min, P >0.05), and so was the time of gastrointestinal function recovery (ï¼»1.27 ± 0.26ï¼½ vs ï¼»2.43 ± 0.41ï¼½ d, P <0.05). No statistically significant differences were observed between the two groups during the 6- to 18-month follow-up in the rates of postoperative subcutaneous hematoma, scrotal and subcutaneous emphysema, recurrence, or pregnancy (P >0.05). CONCLUSIONS: Laparoscopic extraperitoneal high ligation of the spermatic vein is safe and effective and has the advantage of quick recovery in the treatment of varicocele.

[Effects of the Rho-kinase inhibitor fasudil on the invasion, migration, and apoptosis of human prostate cancer PC3 and DU145 cells].

OBJECTIVE: To investigate the potential role of the RhoA/Rock signaling pathway in the formation of prostate cancer and the effects of the Rock inhibitor fasudil on the invasion, migration and apoptosis of human prostate cancer cells. METHODS: Human prostate cancer cell lines PC3 and DU145 were treated with fasudil at the concentrations of 5, 10, 20, 40, 80, and 160 µmol/L, respectively, and those as negative controls cultured in the Ham's-F12 medium, all for 24 hours. Then, MTT assay was used to measure the cell inhibition rate and half maximal inhibitory concentration (IC50) value of fasudil, with 1/4 of IC50 as the medication dose for further investigation. The expressions of RhoA, RockⅠ, and RockⅡ proteins in the PC3 and DU145 cells were detected by Western blot and immunohistochemistry, and the invasion, migration and apoptosis of the cells were determined using the Transwell chamber, scratch wound healing assay and flow cytometry. RESULTS: Fasudil inhibited the proliferation of the PC3 cells from (9.29±1.23)% at 5 µmol/L to (81.37±3.97)% at 160 µmol/L and that of DU145 from (7.59±1.54)% to (76.53±2.67)%, both in a dose-dependent manner (P<0.05 ). Significantly fewer PC3 and DU145 cells migrated into the lower compartment in the experimental group (39.2±8.4 and 34.2±6.7) than in the negative control (116.8±9.3 and 112.5±10.8) (P<0.05 ). The wound healing rates of the PC3 and DU145 cells were remarkably lower in the former (ï¼»37.26±1.17ï¼½% and ï¼»32.38±2.73ï¼½%) than in the latter (ï¼»78.12±4.16ï¼½% and ï¼»69.47±6.71ï¼½%) (P<0.05 ). Annexin V-FITC/PI double staining showed markedly increased apoptosis rates of PC3 and DU145 cells treated with fasudil (ï¼»31.88±2.49ï¼½% and ï¼»28.65±2.99ï¼½%) as compared with the negative controls (ï¼»7.51±2.28ï¼½% and ï¼»7.13±1.61ï¼½%) (P<0.05 ). The expressions of RockⅠ and RockⅡ were significantly reduced in the fasudil-treated cells in comparison with those of the control group (P<0.05 ) while that of RhoA showed no significant difference between the two groups (P>0.05 ). CONCLUSIONS: The RhoA/Rock signaling pathway may play an important role in the formation of prostate cancer. Fasudil can significantly inhibit the proliferation, migration, and invasion and promote the apoptosis of human prostate cancer PC3 and DU145 cells by reducing RhoA/Rho kinase activity.

[Effects and complications of five surgical approaches to the treatment of varicocele: A comparative study].

OBJECTIVE: To compare the effects and complications of subinguinal microscopic ligation, laparoscopic transperitoneal varicocelectomy, laparoscopic retroperitoneal varicocelectomy, open retroperitoneal high ligation, and interventional embolotherapy in the treatment of varicocele. METHODS: We conducted a retrospective study that included 632 varicocele patients treated by subinguinal microscopic ligation (group A, n = 79), laparoscopic transperitoneal varicocelectomy (group B, n = 120), laparoscopic retroperitoneal varicocelectomy (group C, n =137), open retroperitoneal high ligation (group D, n = 283), and interventional embolotherapy (group E, n = 13). We compared the baseline and 3-month postoperative semen parameters, postoperative complications, and pregnancy rate among the five groups of patients. RESULTS: The operation time was longer in groups A ([2.02 ± 1.25] h) and E ([2.17 ± 1.02] h) than in the other three groups, while the postoperative hospital stay was the shortest in group E ([1.1 ± 0.1] d). Intestinal injury or incision bleeding occurred intraoperatively in 2 cases in group B and 1 case in group E. Postoperative scrotal edema developed in 3.7, 17, 10, and 19% of the patients in groups A, B, C, and D, respectively, but not in group E. The rate of 1-year recurrence was the lowest in group A (1.6%) and highest in group E (22%). Sperm concentration and the percentages of progressively motile sperm and morphologically normal sperm were improved postoperatively in all the patients (P < 0. 05), but there were no statistically significant differences among the five groups either in the above three parameters or in the postoperative pregnancy rate (P > 0. 05). CONCLUSION: In the surgical treatment of varicocele, laparoscopic retroperitoneal approach involves short operation time and few complications, subinguinal microscopic ligation has the advantages of little injury, rapid recovery, and few complications but requires specialized microsurgical techniques, and interventional embolotherapy leaves no incision scar and needs only local anesthesia and 1-day postoperative hospital stay, which is uitable for those with a contraindication to anesthesia.